Nonalcoholic steatohepatitis (NASH), a common liver disease, progresses to cirrhosis in 15-20% of subjects. It is not known why only some subjects with NASH progress. This gap in knowledge is a barrier to accurate prediction of prognosis and development of targeted therapy to prevent disease progression. The long-term goal is to understand the mechanisms underlying disease progression in NASH. The objective of this application is to determine how dysregulation of the unfolded protein response (UPR), causes disease progression in NASH. The UPR, which is activated in NASH, is triggered by unfolded proteins in the endoplasmic reticulum (ER). The UPR restores homeostasis by several mechanisms including protein degradation by ER enhancing degradative mannosidase- (EDEM). EDEM is transcriptionally activated by spliced X Box protein-1 (sXBP-1) during the UPR. If homeostasis is not achieved, alarm pathways are activated producing inflammation and apoptosis. The central hypothesis to be tested in this proposal is: EDEM mRNA levels fail to increase despite UPR activation due to translational inhibition of sXBP-1 by specific micro RNA (miRNA), mir451 and mir34a, in some patients with NASH. This retards recovery from UPR and drives alarm pathway activation, apoptosis, inflammation and thus disease progression. Guided by strong preliminary data, this hypothesis will be tested by three specific aims: (1) Identify a molecular signature predictive of disease progression in humans with NASH, (2) Evaluate the effects of over-expressing or silencing EDEM on development and progression of NASH in a mouse model of NASH, and (3) Define the mechanisms responsible for low EDEM expression and function in NASH. The first aim will be achieved by a prospective comparison of the rates of disease progression between those with and without failure of EDEM activation in a cohort of subjects with early stage NASH. The second aim will be achieved by either silencing or over- expressing EDEM in a diet-based mouse model of NASH which is unique because it mimics human disease with respect to histology, insulin resistance, metabolomic profile and pattern of UPR. The third aim will be achieved by measuring the effects of over-expression of mir451 and 34a on EDEM expression and function in vitro and antagonizing mir451 and 34a in vivo in the mouse model of NASH. The miRNA will be silenced with novel phosphorodiamidate-morpholino-oligomers. This approach is innovative because it capitalizes on the novel observation that 20% of subjects with NASH fail to generate EDEM and have the greatest activation of alarm pathways and apoptosis. It also explores the role of miRNA in NASH, a novel field of research. The proposed research is significant because it is expected to (a) provide a way to identify subjects with NASH at risk for progression, (b) identify novel targets for therapy, and (c) advance knowledge related to the role of miRNA in hepatic pathobiology. These studies are thus directly in line with the mission of the NIH that pertains to developing knowledge to reduce the burden of human disease. PUBLIC HEALTH RELEVANCE: If the proposed hypothesis is validated, it will provide a molecular signature identifying those who will progress, and an explanation as to why they progress. This will allow both accurate risk stratification for intervention and development of therapies targeting EDEM or miRNA to prevent cirrhosis due to NASH.